Identification of Signaling Pathways Dysregulated during Scrapie Pathogenesis using a Kinomics Approach

Abstract

Prion diseases are a family of chronic lethal neurodegenerative diseases that affect humans and animals. The conversion of the cellular prion protein (PrPC) to abnormal conformations (PrPSc) is accepted to be required for pathogenesis. However, the molecular mechanisms whereby PrP conversion ultimately results in neuronal death remains poorly understood. Reversible protein phosphorylation by protein kinases and phosphatases regulates signal transduction. Dysregulated protein kinases are involved in the pathogenesis of many chronic diseases, including the neurodegenerative Alzheimer's and Parkinson’s diseases. Consequently, protein kinases have become major therapeutic targets. Protein kinases are also involved in prion disease pathogenesis. However, the signaling pathways most critical to pathogenesis are yet unknown. In this thesis, I describe the development of a multiplex Western blot-based kinomics approach to identify signaling pathways dysregulated during prion disease pathogenesis. This approach was validated using an in vitro model of prion pathogenesis, N2a neuroblastoma cells expressing cytoplasmic PrP mutants. The application of the kinomics approach to an in vivo model, mice infected with scrapie, revealed the dysregulation of two signaling pathways involved in neuronal survival or death at different times after infection. The pro-survival CaMK4β/CREB signaling pathway was activated at pre-clinical stages, but returned to mock-infected levels prior to the onset of clinical disease and the activation of pro-death MST1 signaling. My discovery that CaMK4β/CREB and MST1 signaling is dysregulated in mice infected with scrapie expands our understanding of the molecular mechanisms of prion-mediated neuronal death and identifies potential targets to further evaluate the roles of these pathways in prion disease pathogenesis.